University of Hertfordshire

Standard

A tandem affinity purification tag of TGA2 for isolation of interacting proteins in Arabidopsis thaliana. / Stotz, Henrik; Findling, Simone; Nukarinen, Ella; Weckwerth, Wolfram; Mueller, Martin J.; Berger, Susanne.

In: Plant Signaling and Behavior, Vol. 9, No. 10, e972794, 22.12.2014.

Research output: Contribution to journalArticlepeer-review

Harvard

APA

Vancouver

Author

Stotz, Henrik ; Findling, Simone ; Nukarinen, Ella ; Weckwerth, Wolfram ; Mueller, Martin J. ; Berger, Susanne. / A tandem affinity purification tag of TGA2 for isolation of interacting proteins in Arabidopsis thaliana. In: Plant Signaling and Behavior. 2014 ; Vol. 9, No. 10.

Bibtex

@article{7f5992d4b7a94980bc1394f7dd476870,
title = "A tandem affinity purification tag of TGA2 for isolation of interacting proteins in Arabidopsis thaliana",
abstract = "Tandem affinity purification (TAP) tagging provides a powerful tool for isolating interacting proteins in vivo. TAP-tag purification offers particular advantages for the identification of stimulus-induced protein interactions. Type II bZIP transcription factors (TGA2, TGA5 and TGA6) play key roles in pathways that control salicylic acid, ethylene, xenobiotic and reactive oxylipin signaling. Although proteins interacting with these transcription factors have been identified through genetic and yeast 2-hybrid screening, others are still elusive. We have therefore generated a C-terminal TAP-tag of TGA2 to isolate additional proteins that interact with this transcription factor. Three lines most highly expressing TAP-tagged TGA2 were functional in that they partially complemented reactive oxylipin-responsive gene expression in a tga2 tga5 tga6 triple mutant. TAP-tagged TGA2 in the most strongly overexpressing line was proteolytically less stable than in the other 2 lines. Only this overexpressing line could be used in a 2-step purification process, resulting in isolation of co-purifying bands of larger molecular weight than TGA2. TAP-tagged TGA2 was used to pull down NPR1, a protein known to interact with this transcription factor. Mass spectrometry was used to identify peptides that co-purified with TAP-tagged TGA2. Having generated this TGA2 TAP-tag line will therefore be an asset to researchers interested in stimulus-induced signal transduction processes.",
keywords = "12-oxo-phytodienoic acid, lutathione-S-transferase, lipid stress, protein complex, thale cress",
author = "Henrik Stotz and Simone Findling and Ella Nukarinen and Wolfram Weckwerth and Mueller, {Martin J.} and Susanne Berger",
note = "This is an Accepted Manuscript of an article published by Taylor & Francis Group in Plant Signaling & Behavior, on 31 October 2014, available online: https://doi.org/10.4161/15592316.2014.972794. ",
year = "2014",
month = dec,
day = "22",
doi = "10.4161/15592316.2014.972794",
language = "English",
volume = "9",
journal = "Plant Signaling and Behavior",
issn = "1559-2316",
publisher = "Landes Bioscience",
number = "10",

}

RIS

TY - JOUR

T1 - A tandem affinity purification tag of TGA2 for isolation of interacting proteins in Arabidopsis thaliana

AU - Stotz, Henrik

AU - Findling, Simone

AU - Nukarinen, Ella

AU - Weckwerth, Wolfram

AU - Mueller, Martin J.

AU - Berger, Susanne

N1 - This is an Accepted Manuscript of an article published by Taylor & Francis Group in Plant Signaling & Behavior, on 31 October 2014, available online: https://doi.org/10.4161/15592316.2014.972794.

PY - 2014/12/22

Y1 - 2014/12/22

N2 - Tandem affinity purification (TAP) tagging provides a powerful tool for isolating interacting proteins in vivo. TAP-tag purification offers particular advantages for the identification of stimulus-induced protein interactions. Type II bZIP transcription factors (TGA2, TGA5 and TGA6) play key roles in pathways that control salicylic acid, ethylene, xenobiotic and reactive oxylipin signaling. Although proteins interacting with these transcription factors have been identified through genetic and yeast 2-hybrid screening, others are still elusive. We have therefore generated a C-terminal TAP-tag of TGA2 to isolate additional proteins that interact with this transcription factor. Three lines most highly expressing TAP-tagged TGA2 were functional in that they partially complemented reactive oxylipin-responsive gene expression in a tga2 tga5 tga6 triple mutant. TAP-tagged TGA2 in the most strongly overexpressing line was proteolytically less stable than in the other 2 lines. Only this overexpressing line could be used in a 2-step purification process, resulting in isolation of co-purifying bands of larger molecular weight than TGA2. TAP-tagged TGA2 was used to pull down NPR1, a protein known to interact with this transcription factor. Mass spectrometry was used to identify peptides that co-purified with TAP-tagged TGA2. Having generated this TGA2 TAP-tag line will therefore be an asset to researchers interested in stimulus-induced signal transduction processes.

AB - Tandem affinity purification (TAP) tagging provides a powerful tool for isolating interacting proteins in vivo. TAP-tag purification offers particular advantages for the identification of stimulus-induced protein interactions. Type II bZIP transcription factors (TGA2, TGA5 and TGA6) play key roles in pathways that control salicylic acid, ethylene, xenobiotic and reactive oxylipin signaling. Although proteins interacting with these transcription factors have been identified through genetic and yeast 2-hybrid screening, others are still elusive. We have therefore generated a C-terminal TAP-tag of TGA2 to isolate additional proteins that interact with this transcription factor. Three lines most highly expressing TAP-tagged TGA2 were functional in that they partially complemented reactive oxylipin-responsive gene expression in a tga2 tga5 tga6 triple mutant. TAP-tagged TGA2 in the most strongly overexpressing line was proteolytically less stable than in the other 2 lines. Only this overexpressing line could be used in a 2-step purification process, resulting in isolation of co-purifying bands of larger molecular weight than TGA2. TAP-tagged TGA2 was used to pull down NPR1, a protein known to interact with this transcription factor. Mass spectrometry was used to identify peptides that co-purified with TAP-tagged TGA2. Having generated this TGA2 TAP-tag line will therefore be an asset to researchers interested in stimulus-induced signal transduction processes.

KW - 12-oxo-phytodienoic acid

KW - lutathione-S-transferase

KW - lipid stress

KW - protein complex

KW - thale cress

U2 - 10.4161/15592316.2014.972794

DO - 10.4161/15592316.2014.972794

M3 - Article

C2 - 25482810

VL - 9

JO - Plant Signaling and Behavior

JF - Plant Signaling and Behavior

SN - 1559-2316

IS - 10

M1 - e972794

ER -